The utility of many biologic drugs is limited by inefficient cellular delivery [1]. Previous efforts to overcome this limitation have focused largely on the use of cationic domains, including peptidic cationic species (e.g., HIV-TAT, penetratin, and nonaarginine and more generally cell penetrating peptides (CPP), which are also called protein transduction domains (PTDs)) or non-peptidic cationic species (e.g., PAMAM dendrimers and polyethylenimine), to enhance the attraction between a chemotherapeutic agent and the anionic cell surface [2]. Natural ligands (e.g., folic acid, substance P, and the RGD tripeptide) have also been used to facilitate cellular delivery by targeting agents to specific cell-surface receptors [3]. Such methods have been applied, for example, to delivery of peptides, proteins, nucleic acids and analogs thereof, reporters and labels, various pharmaceuticals and drugs and various small molecules as well as particles. Although some of these methods have had some success, there remains a need in the art for additional delivery strategies. There is a particular need for methods and reagents which facilitate delivery of biologic drugs, particularly peptides and proteins, to the cytosol of cells.
Only a small percentage (<10%) of the extracellular biologics, such as peptides and proteins, that enter endosomes ever get to the cytosol. There is little value in entering endosomes without entering the cytosol.
There are no known methods for modifying a peptide or protein (which does not naturally enter a cell, such as a CPP) to enable its efficient uptake into the cytosol of cells in a bioreversible manner. Being able to do so has many implications for biological research as well as the clinic. For example, dysfunctional proteins could be replaced with functional ones, and misbehaving proteins could be antagonized with specific antibodies. The targeting of antibodies to the cytoplasm is of particular interest [4].